SEARCH · PubMed Health

Results for “Translational repression”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 271 records · Page 15Linked to original sources

PubMed1997

MS2 coat protein mutants which bind Qbeta RNA.

The coat proteins of the RNA phages MS2 and Qbetaare structurally homologous, yet they specifically bind different RNA structures. In an effort to identify the basis of RNA binding specificity we sought to isolate mutants that convert MS2 coat protein to the RNA binding specificity of Qbeta. A library of mutations was created which selectively substitutes amino acids within the RNA binding site. Genetic selection for the ability to repress translation from the Qbetatranslational operator led to the isolation of several MS2 mutants that acquired binding activity for QbetaRNA. Some of these also had reduced abilities to repress translation from the MS2 translational operator. These changes in RNA binding specificity were the results of substitutions of amino acid residues 87 and 89. Additional codon- directed mutagenesis experiments confirmed earlier results showing that the identity of Asn87 is important for specific binding of MS2 RNA. Glu89, on the other hand, is not required for recognition of MS2 RNA, but prevents binding of QbetaRNA.

Allolevivirus↗

PubMed2006

microRNA-mediated silencing inside P-bodies.

Cytoplasmic processing bodies, or P-bodies, contain a high concentration of enzymes and factors required for mRNA turnover and translational repression. Recent studies provide evidence that the mRNAs silenced by miRNAs are localized to P-bodies for storage or degradation, perhaps in adjacent subcompartments. mRNP remodeling, potentially induced by miRISC or RNA helicase activity, may cause the modification of the translation initiation complex at the 5' end of mRNA, following translational repression and localization to P-bodies. Further remodeling in P-bodies may facilitate access of the decapping complex to the cap structure, thus inducing mRNA degradation. However, with appropriate signals, stored mRNAs in P-bodies could be released and returned to the translational machinery through mechanisms requiring binding of regulatory proteins to the 3' UTR of mRNAs. Here a model is proposed to explain the repression and degradation stages of the mRNAs within PBs. This model includes preservation or disruption of a stable closed loop structure of the mRNAs, compartmentalization in PBs and mRNA escape triggered by additional binding proteins.

Cell Compartmentation↗

PubMed2005

Neuronal homeostasis through translational control.

Translational repression is a key component of the mechanism that establishes segment polarity during early embryonic development in the fruitfly Drosophila melanogaster. Two proteins, Pumilio (Pum) and Nanos, block the translation of hunchback messenger RNA in only the posterior segments, thereby promoting an abdominal fate. More recent studies focusing on postembryonic neuronal function have shown that Pum is also integral to numerous mechanisms that allow neurons to adapt to the changing requirements placed on them in a dynamic nervous system. These mechanisms include those contributing to dendritic structure, synaptic growth, neuronal excitability, and formation of long-term memory. This article describes these new studies and highlights the role of translational repression in regulation of neuronal processes that compensate for change.

Animals↗

PubMed1995

Temporal regulation of the Xenopus FGF receptor in development: a translation inhibitory element in the 3' untranslated region.

Early frog embryogenesis depends on a maternal pool of mRNA to execute critical intercellular signalling events. FGF receptor-1, which is required for normal development, is stored as a stable, untranslated maternal mRNA transcript in the fully grown immature oocyte, but is translationally activated at meiotic maturation. We have identified a short cis-acting element in the FGF receptor 3' untranslated region that inhibits translation of synthetic mRNA. This inhibitory element is sufficient to inhibit translation of heterologous reporter mRNA in the immature oocyte without changing RNA stability. Deletion of the poly(A) tract or polyadenylation signal sequences does not affect translational inhibition by this element. At meiotic maturation, we observe the reversal of translational repression mediated by the inhibitory element, mimicking that seen with endogenous maternal FGF receptor mRNA at meiosis. In addition, the activation of synthetic transcripts at maturation does not appear to require poly(A) lengthening. We also show that an oocyte cytoplasmic protein specifically binds the 3' inhibitory element, suggesting that translational repression of Xenopus FGF receptor-1 maternal mRNA in the oocytes is mediated by RNA-protein interactions. These data describe a mechanism of translational control that appears to be independent of poly(A) changes.

Animals↗

PubMed2002

Gene silencing using micro-RNA designed hairpins.

During RNA interference (RNAi), long dsRNA is processed to approximately 21 nt duplexes, short interfering RNAs (siRNAs), which silence genes through a mRNA degradation pathway. Small temporal RNAs (stRNAs) and micro-RNAs (miRNAs) are approximately 21 nt RNAs that are processed from endogenously encoded hairpin-structured precursors, and function to silence genes via translational repression. Here we report that synthetic hairpin RNAs that mimic siRNAs and miRNA precursor molecules can target a gene for silencing, and the mechanism of silencing appears to be through mRNA degradation and not translational repression. The sequence and structural configuration of these RNAs are important, and even slight modification in structure can affect the silencing activity of the hairpins. Furthermore, these RNAs are active when expressed by DNA vectors containing polymerase III promoters, opening the possibility for new approaches in stable RNAi-based loss of function studies.

Blotting, Northern↗

PubMed2002

Rapamycin induces binding activity to the terminal oligopyrimidine tract of ribosomal protein mRNA in rats.

The immunosuppressant rapamycin selectively suppresses the translation of mRNAs containing a terminal oligopyrimidine (TOP) tract adjacent to the cap structure. trans-Acting factors that bind to the 5'-untranslated region (5'-UTR) of TOP mRNAs may be involved in selective translational repression. Some of these factors are regulated by rapamycin-responsive signaling pathways. To identify candidates for the selective trans-acting factor, we examined whether administration of rapamycin alters the binding activity of proteins that bind to RNA containing the TOP element of mouse ribosomal protein (r-protein) L32 mRNA. Preadministration with Freund's complete adjuvant (FCA) prior to rapamycin treatment resulted in increased translational efficiency of r-protein L32 mRNA in submaxillary lymph node (SLN; 2.3-fold), thymus (1.5-fold), and parotid gland (PG; 1.6-fold). Translation of r-protein L32 or elongation factor 1A mRNAs in SLN and PG from FCA-pretreated rats were sensitive to rapamycin administration and the binding ability of p56 was generally increased in extracts from these tissues. On the other hand, in thymus, rapamycin had no effect on the translational efficiency of TOP mRNAs and no p56 binding was detected in the extracts from FCA-pretreated animals. Coadministration of FK506, another immunosuppressive macrolide, increased the p56 TOP-RNA-binding activity and induced selective translational repression of TOP mRNAs in a dose-dependent manner, even in thymus. These findings indicate that p56 is a plausible candidate for the trans-acting factor responsible for regulating the translation of TOP mRNA by a rapamycin-sensitive pathway and that TOP mRNA translational regulation may be responsible for the tissue specificity of rapamycin.

5' Untranslated Regions↗

PubMed2005

MicroRNA-dependent localization of targeted mRNAs to mammalian P-bodies.

Small RNAs, including small interfering RNAs (siRNAs) and microRNAs (miRNAs) can silence target genes through several different effector mechanisms. Whereas siRNA-directed mRNA cleavage is increasingly understood, the mechanisms by which miRNAs repress protein synthesis are obscure. Recent studies have revealed the existence of specific cytoplasmic foci, referred to herein as processing bodies (P-bodies), which contain untranslated mRNAs and can serve as sites of mRNA degradation. Here we demonstrate that Argonaute proteins--the signature components of the RNA interference (RNAi) effector complex, RISC--localize to mammalian P-bodies. Moreover, reporter mRNAs that are targeted for translational repression by endogenous or exogenous miRNAs become concentrated in P-bodies in a miRNA-dependent manner. These results provide a link between miRNA function and mammalian P-bodies and suggest that translation repression by RISC delivers mRNAs to P-bodies, either as a cause or as a consequence of inhibiting protein synthesis.

3' Untranslated Regions↗

PubMed1996

Transcription and masking of mRNA in germ cells: involvement of Y-box proteins.

Gametogenesis is directed by various specialized genetic mechanisms which, to a considerable extent, apply to the production of both eggs and sperm and have been conserved across a wide spectrum of eukaryotic organisms. Two key aspects which are discussed here are: germ-cell-specific gene transcription; and translational repression (masking) of mRNA accumulated in oocytes and spermatocytes/spermatids. Together, these two processes conspire to deliver often large amounts of essential proteins at the appropriate stages of development. It is perhaps not surprising that recent evidence points to a functional link between transcription activation and translation repression, both processes being determined in the nucleus and involving common components. One set of components which has been studied recently are members of the Y-box family of regulatory proteins. Most information of the involvement of Y-box proteins in germ cell development comes from studies on amphibian oocytes and mammalian spermatids. In these cells, Y-box proteins have been detected as major components of both maternal and paternal mRNP particles and have been shown to be instrumental in the masking process. Y-box proteins are also implicated in the regulation of several germ-cell-specific genes. Possible connections between these processes are discussed.

Amino Acid Sequence↗

PubMed1997

Isolation, characterization, and functional studies of rat liver iron regulatory protein 1.

Ferritin mRNAs are translationally regulated by the binding of either of two cytosolic proteins, iron regulatory protein 1 (IRP1) or IRP2, to the iron responsive element (IRE) located in their 5' untranslated region (UTR). Rat liver IRP1 was purified by anion exchange, gel filtration, and affinity chromatography using a concatemerized version of the IRE. Two bands with M(r) of 95,000 and 100,000 were observed by reducing SDS-PAGE. A single protein was responsible for both bands since: (1) [32P]IRE RNA specifically cross-linked to both components; (2) alkylation with iodoacetamide resulted in formation of a single species with M(r) of 95,000; and (3) they possessed identical peptide patterns after digestion with cyanogen bromide. The N-terminal sequence of rat liver IRP1 was MKNPFAHLAEPLDPAQPGKKFNLNKLEDSRYGRLPFXIRVLLEAAV which is identical to the sequence deduced from the cDNA. Rat liver IRP1 has an amino acid composition similar to that of bovine liver caconitase. Several species of IRP1 were observed by two-dimensional gel electrophoresis with pIs ranging from 7.5 to 8.0. Rat liver IRP1 bound the IRE with high affinity (K(D) = 0.04 nM) and repressed translation of ferritin mRNA in vitro. IRP1 bound 100-fold less well to an IRE variant and failed to significantly repress translation of a ferritin mRNA containing the mutated IRE. We conclude that decreases in the affinity of interaction between IRP1 and the IRE, of a magnitude similar to that observed when the binding protein in converted to c-aconitase, are sufficient to significantly enhance translation of ferritin mRNA in vitro.

Amino Acid Sequence↗

PubMed1995

Spnr, a murine RNA-binding protein that is localized to cytoplasmic microtubules.

Previous studies in transgenic mice have established the importance of the 3' untranslated region (UTR) of the spermatid-specific protamine-1 (Prm-1) mRNA in its translational control during male germ cell development. To clone genes that mediate the translational repression or activation of the Prm-1 mRNA, we screened cDNA expression libraries made with RNA from pachytene spermatocytes and round spermatids, with an RNA probe corresponding to the 3' UTR of Prm-1. We obtained six independent clones that encode Spnr, a spermatid perinuclear RNA-binding protein. Spnr is a 71-kD protein that contains two previously described RNA binding domains. The Spnr mRNA is expressed at high levels in the testis, ovary, and brain, and is present in multiple forms in those tissues. Immunolocalization of the Spnr protein within the testis shows that it is expressed exclusively in postmeiotic germ cells and that it is localized to the manchette, a spermatid-specific microtubular array. Although the Spnr protein is expressed too late to be directly involved in the translational repression of Prm-1 specifically, we suggest that the Spnr protein may be involved in other aspects of spermatid RNA metabolism, such as RNA transport or translational activation.

Amino Acid Sequence↗

PubMed1994

Altering the RNA binding specificity of a translational repressor.

The coat proteins of RNA phages MS2 and GA are specific RNA-binding proteins which function to encapsidate viral RNA and to translationally repress synthesis of the viral replicase. The two proteins have highly homologous amino acid sequences, yet they show different RNA binding specificities, recognizing RNA stem-loop structures which differ primarily in the nucleotide sequences of their loops. We sought to convert MS2 coat protein to the RNA binding specificity of GA through the introduction of GA-like amino acid substitutions into the MS2 coat protein RNA-binding site. The effects of the mutations were determined by measuring the affinity of the coat protein variants for RNA in vitro and by measuring translational repression in vivo. We found five substitutions that affect RNA binding. One dramatically reduces binding of MS2 coat protein to both operators. Three others compensate for this defect by nonspecifically strengthening the interaction. Another substitution accounts for the ability to recognize the differences in the RNA loop sequence.

Base Sequence↗

PubMed1999

hnRNP A2 and hnRNP L bind the 3'UTR of glucose transporter 1 mRNA and exist as a complex in vivo.

Recent work identified an RNA binding protein whose presence in brain tumors correlated with translational repression of Glut1 expression. RNase T1 mapping demonstrated that this protein bound an AU-rich response element (AURE) in the Glut1 3'UTR. Facilitated by its differential expression in brain tumor cytosols, we identified this Glut1 RNA binding protein as hnRNP A2. Studies further demonstrated that hnRNP A2 was the major Glut1 RNA binding activity in other cell lines. Recombinant hnRNP A2 exhibited equivalent Glut1 RNA binding specificity, quite distinct from the related AURE binding protein hnRNP A1. These data indicate that hnRNP A2 is the Glut1 AURE binding protein whose cytoplasmic expression in gliomas is associated with translational repression and mRNA instability. Using this approach, we also identified the other major Glut1 3'UTR RNA binding activity as hnRNP L. Stimuli (hypoxia and hypoglycemia) which increase Glut1 mRNA stability selectively decreased polysomal levels of hnRNP A2 and L. Immunoprecipitation demonstrated that hnRNP A2 and L exist as a complex in vivo. As a result of these and other studies, we conclude that hnRNP A2 and L associate in vivo and independently bind the 3'UTR of Glut1 mRNA.

3' Untranslated Regions↗

PubMed2000

Preferential binding of fd gene 5 protein to tetraplex nucleic acid structures.

The gene 5 protein of filamentous bacteriophage fd is a single-stranded DNA-binding protein that binds non-specifically to all single-stranded nucleic acid sequences, but in addition is capable of specific binding to the sequence d(GT(5)G(4)CT(4)C) and the RNA equivalent r(GU(5)G(4)CU(4)C), the latter interaction being important for translational repression. We show that this sequence preference arises from the formation of a tetraplex structure held together by a central block of G-quartets, the structure of which persists in the complex with gene 5 protein. Binding of gene 5 protein to the tetraplex leads to formation of a approximately 170 kDa nucleoprotein complex consisting of four oligonucleotide strands and eight gene 5 protein dimers, with a radius of gyration of 45 A and an overall maximum dimension of 120-130 A. A model of the complex is presented that is consistent with the data obtained. It is proposed that the G-quartet may act as a nucleation site for binding gene 5 protein to adjacent single-stranded regions, suggesting a novel mechanism for translational repression.

Chromatography, Gel↗

PubMed2000

FOG-2, a novel F-box containing protein, associates with the GLD-1 RNA binding protein and directs male sex determination in the C. elegans hermaphrodite germline.

Male sex determination in the Caenorhabditis elegans hermaphrodite germline requires translational repression of tra-2 mRNA by the GLD-1 RNA binding protein. We cloned fog-2 by finding that its gene product physically interacts with GLD-1, forming a FOG-2/GLD-1/tra-2 3'untranslated region ternary complex. FOG-2 has an N-terminal F-box and a novel C-terminal domain called FTH. Canonical F-box proteins act as bridging components of the SCF ubiquitin ligase complex; the N-terminal F-box binds a Skp1 homolog, recruiting ubiquination machinery, while a C-terminal protein-protein interaction domain binds a specific substrate for degradation. However, since both fog-2 and gld-1 are necessary for spermatogenesis, FOG-2 cannot target GLD-1 for ubiquitin-mediated degradation. We propose that FOG-2 also acts as a bridge, bringing GLD-1 bound to tra-2 mRNA into a multiprotein translational repression complex, thus representing a novel function for an F-box protein. fog-2 is a member of a large, apparently rapidly evolving, C. elegans gene family that has expanded, in part, by local duplications; fog-2 related genes have not been found outside nematodes. fog-2 may have arisen during evolution of self-fertile hermaphroditism from an ancestral female/male species.

3' Untranslated Regions↗

PubMed1993

The RNA binding site of bacteriophage MS2 coat protein.

The coat protein of the RNA bacteriophage MS2 binds a specific stem-loop structure in viral RNA to accomplish encapsidation of the genome and translational repression of replicase synthesis. In order to identify the structural components of coat protein required for its RNA binding function, a series of repressor-defective mutants has been isolated. To ensure that the repressor defects were due to substitution of binding site residues, the mutant coat proteins were screened for retention of the ability to form virus-like particles. Since virus assembly presumably requires native structure, this approach eliminated mutants whose repressor defects were secondary consequences of protein folding or stability defects. Each of the variant coat proteins was purified and its ability to bind operator RNA in vitro was measured. DNA sequence analysis identified the nucleotide and amino acid substitutions responsible for reduced RNA binding affinity. Localization of the substituted sites in the three-dimensional structure of coat protein reveals that amino acid residues on three adjacent strands of the coat protein beta-sheet are required for translational repression and RNA binding. The sidechains of the affected residues form a contiguous patch on the interior surface of the viral coat.

Amino Acid Sequence↗

PubMed1996

The role of thymidylate synthase in cellular regulation.

Thymidylate synthase plays a central role in the biosynthesis of thymidylate, an essential precursor for DNA biosynthesis. In addition to its role in catalysis and cellular metabolism, studies from our laboratory have shown that thymidylate synthase functions as an RNA binding protein. Specifically, thymidylate synthase binds with high affinity to its own mRNA resulting in translational repression. An extensive series of experiments have now been performed to elucidate the molecular elements underlying the interaction between thymidylate synthase and its own mRNA. These studies have shed new light into the critical nucleotide sequences and/or secondary structure that are important for protein recognition. As well, studies to define the domains on the protein essential for RNA binding are currently underway. In addition to the characterization of the cis- and trans-acting elements underlying the interaction between thymidylate synthase and its own mRNA, we have recently shown that thymidylate synthase has the capacity to specifically bind in vitro and in vivo to other cellular RNA species. In this regard, thymidylate synthase interacts with the mRNAs of the c-myc onocogene and the p53 tumor suppressor gene. These two genes have been shown to play critical roles in cell cycle control, DNA biosynthesis, and apoptosis. In vitro studies reveal that the interaction of TS with these cell-cycle related mRNAs results in their translational repression. While the biological significance of these cellular RNA/TS protein interactions remains to be defined, these studies suggest a potential role for TS as a mediator in the coordinate regulation of several critical aspects of cellular metabolism.

Antineoplastic Agents↗

PubMed1996

Mechanisms of translational control in early development.

Oocytes accumulate a dowry of maternal mRNAs in preparation for embryogenesis. These maternal transcripts are kept dormant until late oogenesis or early embryogenesis when their translation is activated. In recent years, three types of translational control acting on maternal mRNAs have emerged: translational activation by cytoplasmic polyadenylation, translational activation by RNA localization, and regulated translational repression. In each case, translational control depends on the binding of trans-acting factors to sequences in the 3' untranslated region (3'UTR). Identification of these trans-acting factors is beginning to shed light on the molecular mechanisms that mediate translational control.

Animals↗

PubMed1998

The Pumilio RNA-binding domain is also a translational regulator.

Posterior patterning in the Drosophila embryo requires the action of Nanos (Nos) and Pumilio (Pum), which collaborate to regulate the translation of maternal hunchback (hb) mRNA. Previous work demonstrated that Pum recognizes sites in the 3' UTR of hb mRNA. In this report, we first define the RNA-binding domain of Pum and then show that residues essential for translational repression are embedded within this domain. We also show that Nos and Pum can repress cap-independent translation from an internal ribosome entry site (IRES) in vivo, suggesting that they act downstream of the initial steps of normal, cap-dependent translation.

Amino Acid Sequence↗